Phytochem Rev (2019) 18:241–272

https://doi.org/10.1007/s11101-018-9591-z (0123456789().,-volV)
(0123456789().,-volV)

Comprehensive review of antimicrobial activities of plant

flavonoids
Ireneusz Górniak . Rafał Bartoszewski . Jarosław Króliczewski

Received: 5 December 2017 / Accepted: 29 September 2018 / Published online: 6 October 2018
Ó The Author(s) 2018

Abstract Flavonoids are one of the largest classes of ability to reverse the antibiotic resistance and enhance
small molecular secondary metabolites produced in action of the current antibiotic drugs. Hence, the
different parts of the plant. They display a wide range development and application of flavonoid-based drugs
of pharmacological and beneficial health effects for could be a promising approach for antibiotic-resistant
humans, which include, among others, antioxidative infections. This review aims to improve our under-
activity, free radical scavenging capacity, coronary standing of the biological and molecular roles of plant
heart disease prevention and antiatherosclerotic, hep- flavonoids, focusing mostly on their antimicrobial
atoprotective, anti-inflammatory, and anticancer activities.
activities. Hence, flavonoids are gaining high attention
from the pharmaceutical and healthcare industries. Keywords Antibiotic resistance  Antibacterial
Notably, plants synthesize flavonoids in response to mechanism  Medicinal plants  Nutraceuticals 
microbial infection, and these compounds have been Secondary metabolites
found to be a potent antimicrobial agent against a wide
range of pathogenic microorganisms in vitro. Antimi-
Abbreviations
crobial action of flavonoids results from their various
AHL N-acyl homo-serine lactones
biological activities, which may not seem very specific
AGs Aminoglycoside antibiotics
at first. There are, however, promising antibacterial
CFU Colony-forming units
flavonoids that are able not only to selectively target
EC Epicatechin
bacterial cells, but also to inhibit virulence factors, as
ECG Epicatechin gallate
well as other forms of microbial threats, e.g. biofilm
EGCG Epigallocatechin gallate
formation. Moreover, some plant flavonoids manifest
EPI Efflux pump inhibitors
FAS-I Fatty acid synthase type I
I. Górniak FAS-II Fatty acid synthase type II
Department of Biophysics, Faculty of Biotechnology, FICI Fractional inhibitory concentration index
University of Wroclaw, Fryderyka Joliot-Curie 14a, MBC Minimum bactericidal concentration
50-383 Wroclaw, Poland MDR Multidrug resistant strains
R. Bartoszewski  J. Króliczewski (&) MIC Minimum inhibitory concentration
Department of Biology and Pharmaceutical Botany, MRSA Methicillin-resistant Staphylococcus aureus
Medical University of Gdansk, Hallera 107, MSSA Methicillin-sensitive Staphylococcus
80-416 Gdansk, Poland aureus
e-mail: jakrol@windowslive.com

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PMF Proton-motive force
ROS Reactive oxygen species
QS Quorum sensing
Introduction
Pathogenic microorganisms have been a danger to the
human race since its genesis, being a major cause of
human morbidity and mortality. Until the discovery of
the first true antibiotic—penicillin—in 1928 and sulfa
drugs in the 1930s, besides the toxic arsenic, the only
means of fighting infectious diseases were plant
extracts of different sorts, though their usage yielded
various results (Dar et al. 2016; Saleem et al. 2010; van
Miert 1994).
Although, for the last 60 years, antibiotics played a
major role in the treatment of infectious diseases
caused by bacteria and fungi, the occurrence of
dangerous and antibiotic-resistant bacteria have been
observed to increase in frequency over the past several
decades. Drug resistance can be executed by multiple
mechanisms; hence overcoming such problem is not
an easy task (Saleem et al. 2010). Reasons for the
emerging antibiotic resistance include the irresponsi-
ble, unfit or too common use of antibiotics in fields,
such as medicine, veterinary, and especially in agri-
culture (Pisteli and Giorgi 2012). Moreover, the
pipeline of new antimicrobial agents is running dry
since the 70 s, while the number of drug-resistant
bacteria has increased (Croft et al. 2007; Shah 2005),
leading some to claim that a post-antibiotic era is
eminent (Appelbaum 2012). Hence, there is a pressing
need for finding new antimicrobial drugs.
The use of plants as medicines has a long history in
the treatment of various diseases and to date,
* 100,000 plant species have been tested for their
medicinal use (Schmidt et al. 2012; Veeresham 2012).
In 2007 WHO estimated that 25% of available drugs
are derived from plants used in folk medicine (Cushnie
et al. 2008). Besides the long-established clinical use,
the plant-derived compounds display good tolerance
and acceptance among patients and seem like a
credible source of antimicrobial compounds. Among
109 new antibacterial drugs, approved in the period
1981–2006, 69% originated from natural products
(Newman 2008). One of the major groups of
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Phytochem Rev (2019) 18:241–272
phytochemicals that has been studied extensively for
their antimicrobial properties are flavonoids (Pisteli
and Giorgi 2012). Flavonoids, being mostly plants
pigments, belong to a wide class of chemical com-
pounds (over 6000 different hydroxylated polyphe-
nols) that carry out important functions in plants,
including attracting insects that pollinate, combating
environmental stresses such as microbial infection,
and regulating the cell growth (Falcone Ferreyra et al.
2012; Kumar and Pandey 2013). Fruits and vegeta-
bles are the main dietary sources of flavonoids for
humans.
The flavonoids are known for their antioxidant,
anti-inflammatory, antiallergic, anticancer, antiviral,
and antifungal properties (Harborne and Williams
2000; Havsteen 1983; Havsteen 2002). However,
since plants synthetize flavonoids in response to
microbial infection (Perumal Samy and Gopalakrish-
nakone 2010), there is a growing interest about the
antibacterial properties of flavonoids and their appli-
cation in the therapy for human diseases.
The therapeutic use of flavonoids is supported by
the successful use of preparations containing these
physiologically active constituents in folk medicine.
For example, Tagetes minuta containing querc-
etagetin-7-arabinzylgalactoside was widely used in
the Argentinean folk medicine for the treatment of
various infectious diseases (Tereschuk et al. 1997).
Flower extracts of Retama raetam (Forssk) contain-
ing, among others, licoflavone C and derrone dis-
played antibacterial activity against Gram-positive
and Gram-negative bacteria (Edziri et al. 2012; Hayet
et al. 2008). Tripleurospermum disciforme (known by
the common name Mayweed), used as a disinfectant
and in the treatment of some diseases in folk medicine
of Iran, contains abundance of flavonoids, including
apigenin, kaempferol, luteolin, quercetin, and their
respective glycosides (Tofighi et al. 2015).
In this review, we have summarized the general
information about flavonoid structure, basic proper-
ties, and their occurrence and discuss their scope of
antimicrobial activity as a possible replacement of
conventional antibiotics. Moreover, we have analyzed
the recently reported flavonoid compounds, which
display potent antimicrobial activities, and have
provided examples of flavonoids that manifest syner-
gistic and additive effects upon combining with other
antibiotic drugs.
Phytochem Rev (2019) 18:241–272
Flavonoids structure and nomenclature
Flavonoids are a class of natural phenolic compounds
that include a C6-C3-C6 carbon framework (phenyl
benzopyran). The basic flavonoid structure consists of
a 2-phenyl-benzo-c-pyrane nucleus comprising two
benzene Rings A and B linked through a heterocyclic
pyran or pyrone Ring C. Depending on the level of
unsaturation and oxidation, flavonoids can be grouped
into various subclasses, such as flavones (Fig. 1),
isoflavones (Fig. 2), flavonols (Fig. 3), flavanols
(otherwise known as catechins, Fig. 4), flavanones
(Fig. 5), flavanonols (Fig. 6), chalcones and dihy-
drochalcones (Fig. 7), aurones and anthocyanidins
(Fig. 8), and others that are not noted for their
antimicrobial activities (Falcone Ferreyra et al.
2012) and are not discussed in this paper.
It should not be disregarded, that huge structural
diversity and wide biological activity of flavonoids
comes from their frequent modifications (Chen et al.
2018). Flavonoid glycosides, as well as their preny-
lated, geranylated, methoxylated and hydroxylated
derivatives vary in structure and mode of antibacterial
action (Cushnie and Lamb 2011). The chemical
structures of flavonoids discussed here are presented
in Figs. 1, 2, 3, 4, 5, 7 and 8 (where these compounds
were grouped accordingly to their chemical classes).
Currently, three approaches are being used in
naming flavonoid compounds, which may cause some
confusion, considering the huge number of new
flavonoids being isolated. The most common approach
is using the trivial name that relates to the subclass to
which the compound belongs to, or the plant from
which it was first extracted from. Another approach is
using the semi-systematic name, where the core of the
name comes from the subclass, for example,
3,5,7,30 ,40 -pentahydroxyflavone. The third method is
naming the flavonoids by their systematic chemical
names, for example, 3,4-dihydro-2-phenyl-2H-1-ben-
zopyran (flavan). Although this method is overcom-
plicated in case of common flavonoids, it is the most
precise approach, and thus superior to other naming
approaches, especially when naming novel com-
pounds (Cushnie and Lamb 2005). One of the
recommendations for the flavonoid nomenclature
was prepared by the IUPAC (Rauter 2013). These
recommendations establish rules for the general
nomenclature of flavonoids, providing examples of
acceptable trivial names, and names derived from
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trivial names, along with semi-systematic and fully
systematic names that follow the published IUPAC
recommendations (International Union of Pure and
Applied Chemistry 1993). However, in this paper, we
decided to use the most common approaches (trivial
and semi-systematic for well-known and novel
flavonoids, respectively), as we feel that they are
sufficient for the purposes of this review.
Physiological roles of plant flavonoids
Flavonoids are present in most of the plants, generally
in all of their organs. As the most abundant secondary
plant metabolites, their quantitative distribution varies
from organ to organ or even plant to plant, depending
on the environment. The composition of flavonoids
varies, depending on the plant’s water and nutrient
availability, intensity of sunlight, type of soil, and the
age of the plant (Havsteen 2002). Nevertheless, plants
of the same taxon tend to produce a similar set of
flavonoids, suggesting that genetic predispositions of
plants are dominant (Havsteen 2002; Nicotra et al.
2010).
Those compounds fulfill variety of functions in
plant organs. Anthocyanins along with other flavo-
noids color flowers and fruits, which attracts pollina-
tors and seed dispersers (reviewed in Narbona et al.
2014; Schiestl and Johnson 2013). In vegetative
organs, anthocyanins and other non-pigmented flavo-
noids, such as flavones and flavonols, may provide
some protective functions against many biotic and
abiotic stressors like herbivores, UV radiation, cold,
heat, drought, and salinity (Anderson et al. 2013;
Falcone Ferreyra et al. 2012; Hatier and Gould 2009).
Moreover, flavonoids take part in energy transfers,
regulation of photosynthesis and morphogenesis,
regulation of growth factors, and sex determination
(Harborne and Baxter 1999).
More importantly, there are reports suggesting that
flavonoids are important antimicrobials in plant life.
To arrest the spread of pathogens, plants possess an
innate immunity that involves different layers of
defense responses and some of these defenses include
biosynthesis of flavonoids (Piasecka et al. 2015). Beck
and Stengel (2016) found that flavonoids are mostly
concentrated along the vascular strands of leaves,
rather than being evenly distributed throughout the
leaf tissue. This is due to the need for the quick
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244 Phytochem Rev (2019) 18:241–272

Fig. 1 Chemical structures of flavones

distribution of flavonoids via vascular strands, which feeding, and possibly pathogen attack response. In
shows the important roles of flavonoids in physiolog- fact, flavonoids serve as phytoalexins categorized as
ical regulation, chemical messaging, deterring of the compounds that protect plants from different types of

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Phytochem Rev (2019) 18:241–272
Fig. 2 Chemical structures of isoflavones
pathogens (Cowan 1999). For example, a flavanone
sakuranetin was found in abundance in rice, where it
combats various pathogens, both bacterial and fungal
(Cho and Lee 2015). Moreover, many classes of
flavonoids have been identified as allelochemicals that
inhibit the growth of microorganisms around the plant.
Examples of those include chalcones, dihydrochal-
cones, flavonols, flavanols, flavanones and isoflavo-
noids (Beck and Stengel 2016; Iwashina 2003).
Mechanisms of antimicrobial action by flavonoids
To date, many flavonoids were characterized by the
antibacterial activities against plant pathogens, which
could be effectively applied to fight human pathogens.
Moreover, the antibacterial activities of many plant-
derived flavonoids use different mechanisms than
those of conventional drugs, and thus could be of
importance in the enhancement of antibacterial ther-
apy (Pandey and Kumar 2013).
245
Membrane disruption
The bacterial plasma membrane is responsible for
osmoregulation, respiration and transport processes,
biosynthesis and cross-linking of peptidoglycan, as
well as biosynthesis of lipids. For performing all of
these functions, membrane integrity is a prerequisite,
and its disruption can directly or indirectly cause
metabolic dysfunction and finally lead to bacterial
death (Hartmann et al. 2010). To date, flavonoids,
especially catechins, have been widely studied for
their antimicrobial properties in both Gram-positive
and Gram-negative bacteria. The interactions of
flavonoids with lipid bilayers involve two mechanisms
(Tsuchiya 2015). The first is associated with the
partition of the more non-polar compounds in the
hydrophobic interior of the membrane, while the
second one includes the formation of hydrogen bonds
between the polar head groups of lipids and the more
hydrophilic flavonoids at the membrane interface.
Moreover, nonspecific interactions of flavonoids with
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246 Phytochem Rev (2019) 18:241–272

Fig. 3 Chemical structures of flavonols

phospholipids can induce structural changes in the influence pharmacological properties of flavonoids
properties of the membrane (e.g., thickness and themselves (Arora et al. 2000). However, the mech-
fluctuations) and indirectly modulate the distribu- anism responsible for the flavonoid–membrane inter-
tion/function of membrane proteins, as well as action has not yet been fully understood and the

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Phytochem Rev (2019) 18:241–272
Fig. 4 Chemical structures of
flavanols (catechins)
literature so far remains controversial (Sanver et al.
2016).
Catechins (Fig. 4) are often linked to the antimi-
crobial effects and related to the interactions with the
cell membrane. Contrasting to the protective effects of
flavonoids on membranes, catechins were shown to
247
rupture the bacterial membrane by binding to the lipid
bilayer and by inactivating or inhibiting the synthesis
of intracellular and extracellular enzymes (Reygaert
2014). Moreover, recent studies employing cell mod-
els have highlighted the pro-oxidative activity of
several polyphenols already known as antioxidants,
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248
Fig. 5 Chemical structures of flavanones
namely epicatechin (EC, compound 39), epigallocat-
echin gallate (EGCG, compound 42) and a flavonol
quercetin (32) (Bouayed and Bohn 2010). Fathima and
Rao (2016) reported that the mode of action of killing
bacteria by catechins was found to be an oxidative
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Phytochem Rev (2019) 18:241–272
burst by the generation of reactive oxygen species
(ROS) that cause alteration in the membrane perme-
ability and membrane damage. It should be noted
however, that oxidative bursts occur only at high
EGCG concentrations. Liposome studies also showed
Phytochem Rev (2019) 18:241–272
Fig. 6 Chemical structures of flavanonols
membrane disruption by this compound (Sirk et al.
2009). Interestingly, liposomes containing high
amounts of negatively charged lipids, were less
susceptible to catechin damage, just as catechins have
less effect on Gram-negative bacteria due to nega-
tively charged LPS of the outer bacterial membrane
(Ikigai et al. 1993). It correlates well with studies
reporting lower antibacterial activities of catechins
against Gram-negative bacteria versus Gram-positive
bacteria (Cushnie et al. 2008). Cushnie et al. (2008)
reported that membrane disruption by catechins
causes potassium leakage in methicillin-resistant Sta-
phylococcus aureus (MRSA) strain, which is the first
indication of a membrane damage in microorganisms
(Lambert and Hammond 1973). They have also
noticed that more lipophilic, acylated to 3-O-oc-
tanoyl-epicatechin (43) yields better results in antibac-
terial studies, than unmodified epicatechin (39). The
249
increased activities are the result of enhanced mem-
brane affinity of their long acyl chains (Matsumoto
et al. 2012).
Other flavonoids are also often reported to possess
membrane-disrupting activities. Sato et al. (1997)
reported that 2,4,60 -trihydroxy-30 -methylchalcone
(62) leads Streptococcus mutans to leak intracellular
substances such as protein and ions. Mirzoeva et al.
(1997) noticed that quercetin (32) from propolis
causes a decrease of proton-motive force in S. aureus
and suggested that increased membrane permeability
contributes to the synergistic activity of propolis with
antibiotics, such as tetracycline and ampicillin (Ste-
panovic et al. 2003). Furthermore, Ollila et al. (2002)
showed that flavones acacetin (9) and apigenin (10), as
well as flavonols morin (26) and rhamnetin (37),
caused destabilization of the membrane structure by
disordering and disorientation of the membrane lipids
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250
Fig. 7 Chemical structures of chalcones. It should be noted that
positions of substituents in semi-systematic names may differ
compared to original reports due to variations in numeration of
the positions of the rings in chalcone structures. We adjusted the
and induced leakage from the vesicle. Tsuchiya and
Iinuma (2000) reported that flavanones naringenin
(51) and sophoraflavanone G (56) have antibacterial
activity against MRSA. They have also noticed that
the antibacterial effect of these flavonoids is caused by
reducing the fluidity in hydrophilic and hydrophobic
regions of the both inner and outer cellular membrane.
Sanver et al. (2016) showed that flavonols quercetin
(32), rutin (quercetin-3-O-rhamnoglucoside, com-
pound 35) and tiliroside (38) decreased the bilayer
thickness, furthermore rutin disrupted the lipid mono-
layer structure. Synthetic lipophilic 3-arylidenefla-
vanones (substituted with various phenolic compound
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Phytochem Rev (2019) 18:241–272
nomenclature of chalcones in this paper, according to the most
common numeration approach (Boumendjel 2003). The chem-
ical structures are consistent with the original reports
at the C-3 position of C Ring) were found to be highly
active against S. aureus, Staphylococcus epidermidis,
and Enterococcus faecalis due to flavonoid-initiated
bacterial cell aggregation that influences the integrity
of membranes and causes biofilm disturbance
(Budzynska et al. 2011).
Concluding, differences in the number and distri-
bution of hydroxyl groups, the polymerization degree,
as well as the presence of a methoxy groups in the C
ring, can influence the type of interactions that occur
between different flavonoids and lipid bilayers (Oteiza
et al. 2005). Moreover, flavonoids lacking hydroxyl
groups on their B Rings are more active against
Phytochem Rev (2019) 18:241–272 251

cells to a surface, which is followed by cells dividing

and developing into mature, three-dimensional bio-
films (Costerton et al. 1995). However, Kragh et al.
(2016) have showed that multi cellular bacteria
composites perform better than single cells during
the biofilm development. Interestingly, there are
reports of flavonoids supporting bacterial aggregation.
Stapleton et al. (2004) observed pseudo multicellular
aggregates of S. aureus after incubation with EGCG
(42) and 3-O-octanoyl-epicatechin (43). Flavonols
have also been reported to cause aggregations of
bacterial cells, particularly galangin (25) (Cushnie
Fig. 8 Chemical structures of other flavonoids mentioned in
this paper
et al. 2007). It should be noted however, that growth of
the bacteria was inhibited after aggregation. Presum-
ably, flavonoids cause bacterial aggregation by their
microbial membranes than those with the –OH groups
partial lysis, which leads to membrane fusion, and
(Chabot et al. 1992). This is due to negative correlation
consequently reduces the active nutrient uptake via a
between the relative hydrophobicity of flavonoids and
smaller membrane area, thus it cannot be stated that
the number of hydroxyl group present. Furthermore,
flavonoids support biofilm formation. On the contrary,
other authors suggest that lipophilic flavonoids which
multiple research teams reported that flavonoids in
are highly hydroxylated can be more disruptive for
fact inhibit biofilms. For example, Awolola et al.
membrane structure (Matijašević et al. 2016; Mishra
(2014) showed a significant antibiofilm activity of
et al. 2009; Sato et al. 1996). It is worth noting that
isovitexin (apigenin-6-C-glycoside 14), EC (39) and
bacterial membrane damage by catechins and other
5,7,40 -trihydroxyflavanol (44) against S. aureus ATCC
flavonoids may also result in an inability of the
29213. Similarly, El-Adawi (2012) observed a
bacteria to secrete toxins (Lee et al. 2011; Shah et al.
55–66% decrease in S. mutans biofilm formation upon
2008).
exposure to 2–15% EC. However, Nyila et al. (2012)
observed that EC from Acacia karroo did not reduce
Biofilm formation
Listeria monocytogenes biofilms.
Quorum sensing, in particular, autoinducer-2-me-
Bacterial biofilm-based infections constitute a signif-
diated cell–cell signaling, was proposed as a signifi-
icant amount of all microbial and chronic infections in
cant regulatory factor for the biofilm production in
animals and humans, as well as in food spoilage
Escherichia coli, Vibrio spp., and Salmonella typhi-
(Abdullahi et al. 2016; Jamal et al. 2018). One of the
murium (Vikram et al. 2010). Interestingly, citrus
crucial features of bacteria growing as biofilms is that
flavonoids, such as apigenin (10), kaempferol (28),
they become from 10 to 1000 times more resistant to
quercetin (32) and naringenin (51) are effective
antimicrobial agents when compared to their plank-
antagonists of cell–cell signaling (Vikram et al.
tonic cells (Kon and Rai 2016). The current medicinal
2010). Furthermore, quercetin (assigned with
approaches to eradicate biofilm bacteria using sys-
antileishmanial and antibacterial activities (Gatto
temic antibiotic treatments are very limited. However,
et al. 2002; Prasad et al. 2014)) was shown to inhibit
antibiofilm phytochemical compounds were shown to
enteroaggregative E. coli EAEC 042 biofilm (Barboza
influence the bacterial biofilm establishment and
et al. 2016). Quercetin inhibited alginate production in
growth as well as the related bacterial adhesion,
a concentration-dependent manner, resulting in the
motility, and quorum sensing (QS). Furthermore,
declination in the adherence during biofilm formation.
these compounds are believed to have a lower
Moreover, this flavonoid inhibited N-acyl homoserine
probability of bacterial resistance occurrence (Borges
lactones (AHL)-mediated QS. Most notably, quercetin
et al. 2013).
upregulates the expression of several iron siderophore
Although the initiation of biofilm formation has
proteins limiting the amount of Fe3? that is required
been thought to be due to random attachment of single
for the biofilm formation of Pseudomonas aeruginosa

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252
(Ouyang et al. 2016; Symeonidis and Marangos 2012).
Kaempferol (28), epicatechin gallate (ECG, com-
pound 40) and EGCG (42) were reported to mediate
the displacement of AHL molecules from LuxR-type
transcriptional activator protein (Roy et al. 2017),
while chrysin (12), phloretin (68) and naringenin (51)
inhibited QS synthase/receptor pairs, LasI/R, and
RhlI/R (Paczkowski et al. 2017). Cranberry A-type
proanthocyanidins (Fig. 8) are also found to be anti-
adhesion agents against the Gram-negative bacterium
P. aeruginosa (Ulrey et al. 2014). Ulrey et al. (2014)
suggested that the mechanism of A-type proantho-
cyanidins against the biofilm formation results from
their chelating properties.
Hydrophilic flavonoids can interact at the mem-
brane surface and provide protective actions against
different deleterious agents and biofilm formation
(Oteiza et al. 2005). However, Lee et al. (2011)
showed that biofilm reduction by flavonoids does not
result from their antioxidant properties alone. They
demonstrated that flavones, such as 6-aminoflavone
(1), 6-hydroxyflavone (2), apigenin, chrysin (12), as
well as isoflavones daidzein (21) and genistein (22),
and a dihydrochalcone phloretin (68) had inhibitory
effects on E. coli O157:H7 biofilm formation,
although antioxidant compounds (vitamin C and
vitamin E) did not show such effect. Furthermore,
phloretin (a natural, nontoxic apple flavonoid) caused
the most significant reduction of enterohemor-
rhagic E. coli O157:H7 biofilms without affecting
the growth of planktonic cells. Similar effect was
showed for the auronol called derriobtusone A (74)
that inhibited the biofilm formation in E. coli, although
the planktonic growth of E. coli was only weakly
inhibited (Vasconcelos et al. 2014). Notably, phloretin
(68) showed a dose-dependent inhibition of biofilm
and did not harm commensal E. coli K-12 and
nonpathogenic E. coli ATCC 4157 biofilm (Lee
et al. 2011). This is an important feature of phloretin,
since antibiofilm agent should be able to selectively
inhibit the pathogenic strains without wiping out the
commensal microflora.
Fimbriae, including curli and pili, are important
factors for the biofilm formation (Rendón et al. 2007).
Phloretin (68) reduced fimbriae formation in E. coli
O157:H7, due to repression of the expression of the
curli genes (csgA and csgB) (Lee et al. 2011). This
study also reported that phloretin repressed the
expression of two toxin genes (hemolysin hlyE and
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Phytochem Rev (2019) 18:241–272
Shiga toxin 2 stx2). However, phloretin was also
shown to induce stress resistance genes, such as
marRAB and hcsBA genes (Lee et al. 2011). Thus,
phloretin could positively influence the antibiotic
resistance as well.
Efflux-pump inhibitors (EPI) are aimed not only to
block the efflux pumps, but also the biofilm formation
(Sana et al. 2015). Pinostrobin (a dietary flavanone
discovered in the wood of pine, Pinus strobus,
compound 53) enhanced membrane permeability in
both Gram-positive and Gram-negative bacteria (E.
faecalis, S. aureus, E. coli and P. aeruginosa), which
correlated well with its effect on EPI and antibiofilm
formation in Gram-negative bacteria (Christena et al.
2015). Christena et al. (2015) suggested that pinos-
trobin exerts its antibiofilm effect by the mechanism
that is unrelated to its EPI effect and may not involve
the repression of curli genes. This is in contrast to the
report of pinostrobin EPI effect in S. typhimurium
(Baugh et al. 2012). Tea EGCG (42) provides another
example of effective antimicrobial agent against both
the planktonic and biofilm forms of E. faecalis. Tea
EGCG inhibits not only the bacterial growth, but also
suppresses the expression of specific genes related to
biofilm formation (Lee and Tan 2015). A number of
prenylated flavonoids isolated from Epimedium spe-
cies inhibited Porphyromonas gingivalis biofilm for-
mation, however, the antibiofilm mechanism of those
flavonoids remains unknown (Kariu et al. 2016;
Olczak et al. 2005).
Inhibition of cell envelope synthesis
Bacterial-type II fatty acid synthase (FAS-II) differs in
many ways from the mammalian one (FAS-I), which
makes it excellent target for an antimicrobial agent.
Multiple inhibitors of the FAS-II components have
been reported to date and summarized below.
Quercetin (32), apigenin (10), and sakuranetin (54)
were shown to inhibit 3-hydroxyacyl-ACP dehydrase
from Helicobacter pylori (Zhang et al. 2008b).
Extensive research has been made on 3-ketoacyl-
ACP synthase from E. faecalis and 11 flavanones with
different configurations of hydroxyl groups have been
screened (Jeong et al. 2009). The best result was
obtained for the use of eriodictyol (49), naringenin
(51) and taxifolin (60). Parallel docking studies,
conducted by the same team, indicate that hydrogen
bonds between flavonoid hydroxyl groups at C-40 and
Phytochem Rev (2019) 18:241–272
C-50 of B ring and enzyme amino acid residues Arg38
and Phe308 were the key for their antibacterial activity
(Figs. 5, 6). Elmasri et al. (2017) reported 5,6,7,40 ,50 -
pentahydroxyflavone (3) and 5-hydroxy-40 ,7-
dimethoxyflavone (5) to downregulate the malonyl
CoA-acyl carrier protein transacylase fabD (MCATs)
that regulates bacterial FAS-II. Thus, these two
flavones are considered to be the promising drugs for
blocking the bacterial growth. Furthermore, EGCG
(42) from green tea inhibited specific reductases
(FabG, FabI) in the bacterial FAS-II (Zhang and Rock
2004). FabG enzyme (beta-ketoacyl-[acyl carrier
protein] reductase) participates in the fatty acid
biosynthesis and is the only known isoenzyme to
catalyze the reduction of the bacterial membrane b-
keto groups (Li et al. 2006). Therefore, this enzyme is
an ideal target for the development of new antibiotics.
Inactivation of FabG probably occurs as a result of
EGCG-induced aggregation of this enzyme. Finally,
other enzymes involved in fatty acid biosynthesis,
such as 3-ketoacyl-ACP reductase and enoyl-ACP
reductase from many bacteria are inhibited by EGCG
as well (Li et al. 2006; Zhang et al. 2008a; Zhang and
Rock 2004).
Mycobacteria cause some of the most serious
diseases, which are notoriously difficult to treat (Chen
et al. 2010). The presence of mycolic acids is one of
the most distinctive and essential survival features of
the mycobacterial cell wall. Those bacteria possess
two types of fatty acid synthases, a mammalian-type
FAS-I, and a bacterial-type FAS-II, both of which are
important for the biosynthesis of mycolic acid. A
number of flavonols have been shown to inhibit FAS-I,
including: quercetin (32), kaempferol (28), fisetin
(24), morin (26) and myricetin (27), as well as flavones
baicalein (11) and luteolin (15), and EGCG (42) (Li
and Tian 2004). Moreover, some of these flavonoids
possessed activity against FAS-II components as well,
including enoyl-ACP-reductase, b-ketoacyl-ACP
reductase, and b-hydroxyacyl-ACP dehydratases
(Brown et al. 2007). Furthermore, Brown et al.
(2007) reported that chalcones 4,20 ,40 -trihydroxychal-
cone (61), butein (64), isoliquirtigenin (65), and a
flavonol fisetin (24) possess inhibitory activity against
FAS-II from Mycobacterium bovis BCG.
Peptidoglycan is an essential component of the
bacterial cell wall, and the inhibition of its synthesis is
a common mechanism of action of conventional
antimicrobial drugs and flavonoids. Baicalein (flavone
253
from Scutellaria baicalensis, 11) supported induced
by EGCG (42) peptidoglycan damage (Fujita et al.
2005). Flavonols galangin (25), kaempferide (30), and
kaempferide-3-O-glucoside (31) showed not only
activity against amoxicillin-resistant E. coli, but also
the ability to reverse the resistance via inhibition of
peptidoglycan and ribosome synthesis (Eumkeb et al.
2012). Another study on the mechanism of action of
catechins showed that they interfere with the biosyn-
thesis of the bacterial cell wall by binding with the
peptidoglycan layer. Cell wall synthesis was also
inhibited by a synergistic effect of EGCG (42) and
DL-cycloserine (an inhibitor unrelated to penicillin-
binding protein) (Zhao et al. 2001). Furthermore, since
both EGCG and b-lactams (benzylpenicillin, oxacil-
lin, methicillin, ampicillin, and cephalexin) directly or
indirectly target peptidoglycan (Zhao et al. 2001),
EGCG synergizes the activity of b-lactams. Kinetic
studies on D-alanine-D-alanine ligase, responsible for
the production of the terminal dipeptide of peptido-
glycan precursor UDPMurNAc-pentapeptide, showed
that quercetin (32) and apigenin (10) inhibit this
enzyme (Wu et al. 2008). These two flavonoids bind to
the active center of D-alanine-D-alanine ligase (Singh
et al. 2013; Wu et al. 2008). However, quercetin had
poorer activity compared to apigenin, which is
attributed to its additional -OH groups that enforce
its affinity to the enzyme (Figs. 1, 3). In the contrast,
sakuranetin (54), a flavonoid similar to apigenin (it has
7-methoxy instead of 7-hydroxy group, and no double
bond on C ring, Figs. 1, 5), has no inhibitory effect
(Wu et al. 2008). Furthermore, the hydrophilic nature
of quercetin limits its penetration into the bacterial
cell.
Inhibition of nucleic acid synthesis
Flavonoids have been reported to be significant
topoisomerases inhibitors, which contributes to their
antimicrobial activity. For example, DNA gyrase is an
essential enzyme for the DNA replication and it is
exclusive to prokaryotes, which makes it an attractive
target for antibacterial drugs (Plaper et al. 2003).
Ohemeng et al. (1993) reported the inhibition of DNA
gyrase from E. coli by quercetin (32), apigenin (10),
and 3,6,7,30 ,40 -pentahydroxyflavone (4). Moreover, in
silico analysis suggested that subunit B of DNA gyrase
from Mycobacterium smegmatis and M. tuberculosis
can be targeted by quercetin (Suriyanarayanan et al.
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2013). This report was confirmed, by the studies
conducted on different gyrase subunits that revealed
quercetin binding to the B subunit of gyrase and the
corresponding blockage of ATP binding pocket by the
formation of hydrogen bonds via 5, 7 and 30 –OH
groups to the amino acid residues of DNA gyrase
(Fig. 3) (Plaper et al. 2003). It is in correlation with the
studies that reported the blockage of ATP binding
pocket of D-alanine-D-alanine ligase by the same
flavonoids (Wu et al. 2008). Moreover, the related
flavonoids chrysin (12) and kaempferol (28) greatly
inhibited DNA gyrase from E. coli (nobiletin (16),
tangeritin (19) and myricetin (27) were less efficient
inhibitors) (Wu et al. 2013). Those studies showed that
flavonoid hydroxyl groups allow better association
with the gyrase compared to methoxy groups,
although an extra 50 -OH in myricetin greatly
decreased its gyrase inhibition properties (Fig. 3)
(Wu et al. 2013). The second mechanism of DNA
gyrase inhibition was proposed by molecular docking
studies (Fang et al. 2016; Plaper et al. 2003), which
suggest that flavonoids inhibit the DNA supercoiling
by competitively interacting with the ATP binding site
of the DNA gyrase B subunit (GyrB). In this mech-
anism of action, flavonoids binding to DNA stabilizes
the DNA–gyrase complex that leads to DNA cleavage
induction (Plaper et al. 2003). Moreover, Fang et al.
(2016) reported 3-hydroxyl, 5-hydroxyl, 7-hydroxyl,
and 4-carbonyl groups to be crucially active sub-
stituents of flavonoids by interacting with key residues
of GyrB. This result is in accordance with previous
studies of Wu et al. (2013). Furthermore, Ulanowska
et al. (2006) showed that isoflavone genistein (22)
inhibits the growth of Vibrio harveyi (with interme-
diate effect on Bacillus subtilis and little effect on
E. coli) in a dose–response manner. They suggested
that the inhibition of growth of the bacteria species
results from genistein-mediated stabilization of the
topoisomerase II–DNA cleavage complex that leads to
the impairment of cell division and/or completion of
chromosome replication (Verdrengh et al. 2004).
Helicases are ubiquitous motor proteins that sepa-
rate and/or rearrange nucleic acid duplexes in reac-
tions fueled by adenosine triphosphate (ATP)
hydrolysis (Shadrick et al. 2013). Similarly to topoi-
somerases and gyrases, their function is essential for
DNA replication. Recent studies suggested these
proteins as molecular targets of flavonoids. Flavones
and flavonols, the groups of pharmacophores with
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nucleic acid binding capacity, have been screened as
helicase inhibitors. A flavone luteolin (15) and its
structurally related flavonols, such as morin (26)
myricetin (27), were shown to inhibit the replicative
helicases like DnaB and RecBCD helicase/nuclease of
E. coli (Xu et al. 2001). Moreover, myricetin inhibited
Gram-negative bacterial growth and was proposed to
be a potent inhibitor of numerous DNA and RNA
polymerases, as well as viral reverse transcriptases
(along with baicalein (11)) (Ono et al. 1990) and
telomerases (Griep et al. 2007).
Dihydrofolate reductase (DHFR) is a common
target of many drugs, including antimicrobial agents.
The DHFR is an important enzyme of the folic acid
synthesis pathway, which provides precursor of
pyrimidines and purines (Bhosle and Chandra 2016).
EGCG (42) was reported to inhibit DHFRs from
Streptomonas maltophilia, Mycobacterium tuberculo-
sis, and E. coli (Navarro-Martinez et al. 2005; Raju
et al. 2015; Spina et al. 2008). Furthermore, EGCG
had synergistic effects with other inhibitors of folic
acid pathway, such as sulfamethoxazole and etham-
butol (Navarro-Martinez et al. 2005; Raju et al. 2015).
Flavonoid DNA intercalation, that inhibits bacterial
nucleic acid synthesis, was also proposed as a
mechanism underlying their antimicrobial properties.
Mori et al. (1987) noticed that the incubation with
EGCG (42), myricetin (27), and robinetin (17) resulted
in reduced DNA, RNA, and protein synthesis by
Proteus vulgaris and S. aureus. They proposed that
this process resulted from the intercalation of
flavonoids with nucleic acids, mediated by flavo-
noid-free hydroxyl group at C-3 of A ring and 30 ,40 ,50 -
trihydroxyl motif at B ring (Figs. 1, 3, 4). However,
DHFR inhibition could explain the reduction of DNA
and RNA synthesis by EGCG, as well. Furthermore,
myricetin and robinetin, which share similar structure,
also seem to be the possible DHFR inhibitors. These
observations raise the question, ‘‘whether those com-
pounds only reduce nucleic acid synthesis via DNA
intercalation/DHFR inhibition or they have got mul-
tiple target sites?’’ Giving the low specificity of
EGCG, lowered DNA and RNA synthesis could result
from multiple enzyme inhibitions and proton-motive
force (PMF) disruption. Numerous studies reported on
flavonoid-mediated topoisomerase inhibition and
DNA intercalation in human cancer cells (reviewed
by Russo et al. (2012)), suggesting the universal
mechanisms of their action.
Phytochem Rev (2019) 18:241–272
Inhibition of electron transport chain and ATP
synthesis
The membrane potential, being the essential main
energy source for almost all chemical processes in
living systems, is the most important factor for the
survival and growth of bacterial cells. Notably, the
treatment of S. aureus with isobavachalcone (66) and
6-prenylapigenin (7) from Dorstenia species resulted
in bacterial membrane depolarization (Dzoyem et al.
2013). Furthermore, Haraguchi et al. (1998) reported
that licochalcones from Glycyrrhiza inflata inhibited
oxygen consumption in Micrococcus luteus cells, and
the site of inhibition was thought to be between CoQ
and cytochrome c in the bacterial electron transport
chain. Although licochalcones A, B, C, and D
(compounds 69–72) caused inhibition of NADH-
cytochrome c reductase activity in the membrane
fraction, while cytochrome c oxidase was not inhib-
ited. However, only licochalcones A and C manifested
antibacterial activities against Gram-positive bacteria
and it was attributed to the presence of lipophilic
prenyl moiety on the D ring of licochalcones A and C
(Fig. 7), which facilitates their infiltration into the
bacterial cell (Haraguchi et al. 1998).
Recently, it has been reported that flavonoids can
inhibit F1FO ATPase of E. coli (Chinnam et al. 2010).
ATP synthase is a highly conserved enzyme with two
sectors, F1 and FO. In E. coli, F1 is composed of
a3b3cdeab2c10, while FO consists of ab2c10. ATP
hydrolysis and synthesis occur on three catalytic sites
in the F1 sector, whereas proton movement occurs
through the membrane-embedded FO (Senior et al.
2002). A wide range of polyphenols has been shown to
bind at the distinct polyphenol binding site and inhibit
the ATP synthase. The polyphenol binding pocket lies
at the interface of a, b, and c-subunits of F1 sector.
Therefore, the proposed mode of flavonoid inhibitory
action was the binding at the polyphenol binding
pocket of ATP synthase and the blockage of clockwise
or anticlockwise rotation of the c-subunit (Gledhill
et al. 2007). Furthermore, the polyphenol binding
pocket residues are highly conserved among different
species including human, bovine, rat, and E. coli
(Walker et al. 2000) Thus, there is great chance that
other microorganisms may be susceptible to this type
of inhibition (Chinnam et al. 2010). The most effective
inhibitors of E. coli F1FO ATPase include baicalein
(11), morin (26), EC (39), as well as flavanonols
255
silibinin (58) and silymarin (59) (Chinnam et al.
2010). Furthermore, quercetin (32), quercetin-3-glu-
coside (isoquercetin, 33) and quercetin-3-O-rham-
noside (quercitrin, 34) are known to prevent the ATP
hydrolysis, although not the ATP synthesis (Chinnam
et al. 2010). EGCG (42) inhibited the acidogenic and
aciduric properties of S. mutans, probably by the
inhibition of the enzymatic activity of F1FO ATPase
(Xu et al. 2011, 2012). Ulrey et al. (2014) demon-
strated that the treatment of P. aeruginosa with A-type
proanthocyanidins (isolated from Cranberries, mono-
mer shown on Fig. 8—compound 73) downregulated
the proteins involved in ATP synthesis: a cytochrome
c (NP_251172), hypothetical protein (NP_251171); as
well as protein subunits of acetyl-CoA carboxylase
(NP_254123), fumarase (NP_253023), and aconitate
hydratase (NP_249485).
A decline in the overall bacterial metabolism can
lead to the indirect arrest of the biofilm formation, as
well. Notably, the 40 ,50 ,5-trihydroxy-6,7-dimethoxy-
flavone (8) (from Teucrium polium) was reported to
affect the F-type ATP synthase (atpD) and thus reduce
the ATP availability in S. aureus (Elmasri et al. 2017).
Antibacterial action of flavonoid-metal complexes
Havsteen (2002), in his voluminous paper on flavo-
noid properties, tried to explain the antibacterial
activities of flavonoids. Since many studies showed
the ability of flavonoids to chelate transition metal ions
(Karlı́čková et al. 2015; Li et al. 2015; Riha et al. 2014;
Samsonowicz et al. 2017), he pointed out that many
flavonoids could cause the inhibition of bacterial metal
enzymes. This mechanism of action is common for
many other antibacterial substances, including lacto-
ferrin from human milk.
Flavonoid chelation sites include two proximal
hydroxyl groups (o-dihydroxyl group in ring B or ring
A), the 3-hydroxy-4-keto group of the C ring or via the
5-hydroxy-4-keto position of the A and C rings.
Although the antibacterial activity of complexes
depends strongly on the metal ion, the preferred metal
binding site depends on the flavonoid, and on the pH
value (Kasprzak et al. 2015). Literature data suggested
that the flavonoids predominantly form complexes
with a metal in 1:2 ratio and that their binding
efficiency is also associated with the transition state of
metal ions (e.g., Fe2? [ Fe3?) (Ren et al. 2008). One
of the well-known flavonoid–metal complexes are the
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quercetin (32) complexes. Bravo and Anacona (2001)
demonstrated that Mn2?, Hg2?, Co2?, and Cd2?
complexes of quercetin show bactericidal effect
against S. aureus, Bacillus cereus, P. aeruginosa,
E. coli, and Klebsiella pneumoniae. Comparatively,
quercetin alone at the same concentration had no
activity. Similar reports are available for morin (26,
Mg2? and Ca2? complexes) against Micrococcus
flavus and S. aureus (Panhwar and Memon 2011)
and 40 ,7-dimethylapigenin (6, Cu2?, Ni2?, Co2?,
Zn2?, Fe3?, Cr3?, Cd2?, and Mn2?) against E. coli,
S. aureus, and P. vulgaris (Wang et al. 1992). Despite
these studies, the antibacterial mechanism of flavo-
noid–metal complexes have not been conclusively
established yet. For example, the La3? and Gd3? (their
metal–ligand ratio was 6:3, and 8:3, respectively)
complexes of morin (26) had lesser antibacterial
activity, when compared to their parent flavonoids
(Kopacz et al. 2005). Complexation with metal ions
causes changes in the flavonoid structure, in their
affinities to various intracellular targets, as well as in
their antioxidant and prooxidant properties. Hence, the
different antibacterial activities of the flavonoid–metal
ion complexes result from their interaction with
different targets than their parent flavonoids. By all
means, an antimicrobial resistance to metals (reviewed
by Hobman and Crossman (2015)) cannot be
excluded.
Inhibition of bacterial toxins
Important virulence factors, such as bacterial hyalur-
onidases (produced by both Gram-positive and Gram-
negative bacteria), directly interact with host tissues or
mask the bacterial surface from host0 s defense mech-
anisms. In the bacterial pathogenesis, hyaluronidase-
mediated degradation of hyaluronan increases the
permeability of connective tissues and decreases the
viscosity of body fluids (Girish and Kemparaju 2007).
Notably, flavonols, such as myricetin (27) and
quercetin (32) have been identified as hyaluronic acid
lyase (Hyal B) inhibitors in Streptococcus agalactiae.
The inhibitory effect of the flavonoids increased with
the number of hydroxyl groups present in the
flavonoid structure (Hertel et al. 2006). However,
hyaluronate lyases from Streptomyces hyalurolyticus
(Hyal S), and Streptococcus equisimilis (Hyal C) were
only inhibited slightly (Hertel et al. 2006).
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Flavonoids, especially catechins and proantho-
cyanidins (due to antioxidant properties), were pro-
posed to neutralize bacterial toxic factors originating
from Vibrio cholerae, S. aureus, Vibrio vulnificus,
Bacillus anthracis, and Clostridium botulinum
(Ahmed et al. 2016; Choi et al. 2007; Delehanty
et al. 2007). Similarly, genistein (22) inhibited the
exotoxin from S. aureus, while kaempferol (28),
kaempferol-3-O-rutinoside (29), and quercetin gly-
coside inhibited the neurotoxin from C. botulinum
(Sawamura et al. 2002). The a-hemolysin (Hla), a
member of bacterial pore-forming b-barrel toxins, is
one of the most important virulence factors produced
by S. aureus. Soromou et al. (2013) reported that
pinocembrin (52), a honey flavanone, reduced S.
aureus a-hemolysin production in a concentration-
dependent manner (pinocembrin reduces the tran-
scription level of Hla and d-haemolysin genes).
Pinocembrin have also been studied to evaluate its
mechanism of actions on the bacterial membranes of
Neisseria gonorrhoeae. Although the pinocembrin-
induced cell lysis has been observed in the study,
mechanisms of actions of this compound have not
been fully elucidated (Rasul et al. 2013; Ruddock et al.
2011). Sugita-Konishi et al. (1999) reported that
EGCG (42) and gallocatechin gallate (GCG, 41)
suppressed the release of verotoxin from enterohem-
orrhagic E. coli cells and concluded that green tea
catechins can be used to prevent the food poisoning
caused by E. coli.
In conclusion, flavonoids manifest many interesting
mechanisms of antibacterial action (Fig. 9). There are
however, antibacterial flavonoids with little known
mechanism, as well as the ones with multiple cellular
targets. Further investigation of action mechanisms
and structure–activity relationship could help us not
only to reveal novel antimicrobials, but also to find the
most target-specific ones, which in case of possible
therapeutic application of flavonoids, remains critical.
Flavonoids as antimicrobial potentiators
Mechanism of resistance to antibacterial agents
Pathogenic bacteria may gain the resistance to antibi-
otic drugs through different mechanisms, such as
prevention of interaction of the drug with the target,
efflux of the antibiotic from the cell, and direct
Phytochem Rev (2019) 18:241–272
Fig. 9 Diagrammatic representation of action mechanism of
flavonoids. Flavonoids can kill or inhibit bacterial cells in
variety of ways, such as causing membrane disruption (1) and
inhibition of nucleic acid synthesis (2a—inhibition of dihydro-
folate reductase (DHFR), 2b—helicase inhibition, 2c—gy-
rase/topoisomerase inhibition), as well as inhibit bacterial
virulence, e.g. toxins (3) and quorum sensing, which impairs
their ability to form biofilms (4). Antimicrobial action can be
also executed through inhibition of cell envelope synthesis,
destruction or modification of the drug compound
(Fig. 10). Moreover, bacteria can share the resistance
genes, for example, the gene of b-lactamase, an
enzyme hydrolyzing the amide bond in the b-lactam
ring through transformation (incorporation of naked
DNA), transduction (phage-mediated), and conjuga-
tion (Fig. 10). Gram-negative bacteria prefer b-lactam
ring hydrolysis, whereas resistance to in Gram-posi-
tive bacteria is mostly achieved by modifications of
the target site of antibiotics (Bush 2013; Bush and
Fisher 2011).
Another bacterial strategy to cope with the presence
of antibiotics is to produce enzymes that inactivate the
drug by adding specific chemical moieties to this
compound. In the case of Gram-negative bacteria, the
aminoglycoside group of antibiotics becomes ineffec-
tive due to the phosphorylation, adenylation, or
acetylation of the antibiotic molecule (Munita and
257
which involves inhibition fatty acid synthase (FAS—5) and
peptidoglycan synthesis (7a—inhibition of Ala–Ala dipeptide
synthesis, 7b—inhibition of peptidoglycan cross-linking).
Flavonoids can inhibit efflux pumps as well, which can lead to
reversing antimicrobial resistance (6). Moreover inhibition of
NADH-cytochrome c reductase activity in the bacterial
respiratory chain (8) and inhibition of ATP synthase (9) were
also reported
Arias 2016). The aminoglycoside-modifying enzymes
(AMEs) that covalently modify the hydroxyl or amino
groups of the aminoglycoside molecule become the
predominant mechanism of aminoglycoside resistance
worldwide (Ramirez and Tolmasky 2010). Further-
more, the chloramphenicol acetyltransferases chemi-
cally inactivate chloramphenicol in both Gram-
positive and Gram-negative bacteria (Schwarz et al.
2004).
Bacteria have also developed mechanisms that
decrease the antibiotic uptake by preventing the
antibiotic from reaching its intracellular or periplas-
mic target. Hydrophilic molecules such as b-lactam
antibiotics, tetracyclines, and some fluoroquinolones
are translocated through the membrane by water-filled
diffusion channels known as porins (i.e., OmpF,
OmpC, and PhoE) (Pages et al. 2008). Bacteria
decrease porin-mediated antibiotic uptake by either a
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258
Fig. 10 Diagrammatic representation of mechanisms of antibi-
otic resistance. Antibiotic resistance can be executed in many
different ways, such as efflux of the antibiotics from the
bacterial cell (1); changing membrane potential, which prevents
antibiotic molecules from entering (2); bypassing target site of
the antibiotic through incorporation of changed precursor (3) or
changing target site by methylation of RNA, mutations, etc. (4).
Antibiotic action can also be abolished through degradation (5)
shift in the type of porins expressed (Domenech-
Sanchez et al. 2003), or by changing the level of porin
expression, as well as by impairing the function of
these channels (Fernández and Hancock 2012). More-
over, the described changes in the membrane perme-
ability are often accompanied by an increased
expression of efflux pumps, in both Gram-negative
and Gram-positive bacteria. The efflux pumps may be
substrate-specific (tetracycline or macrolides, such as
erythromycin in pneumococci) or have broad substrate
specificity, which is common for multidrug resistance
bacteria (MDR) (Poole 2005). There are six major
families of efflux pumps: the ATP-binding cassette
(ABC) superfamily (Lubelski et al. 2007), the major
facilitator superfamily (MFS) (Pao et al. 1998), the
multidrug and toxic compound extrusion (MATE) (Lu
2016), the small multidrug resistance (SMR) family
(Bay et al. 2008) (a member of the much larger drug/
metabolite transporter family (DMT) (Piddock 2006)),
the resistance nodulation division (RND) superfamily
(Nikaido and Takatsuka 2009), and newly discovered
proteobacterial antimicrobial compound efflux pump
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Phytochem Rev (2019) 18:241–272
and chemical modification of antibiotic molecules (6). More-
over, Gram-negative bacteria are resistant to penicillin and other
hydrophilic antibiotics due to the low permeability of their outer
membrane, as well as low expression of porins (7). Furthermore,
bacteria can gain and exchange the resistance genes through
transduction (phage-mediated), conjugation (acquiring plasmid
DNA) and transformation (incorporation of naked DNA) (8)
(PACE) (Hassan et al. 2013). These families differ in
terms of structural conformation, a range of substrates,
energy sources, and in the types of bacterial organisms
in which they are distributed (Soto 2013).
The last common mechanism of antibiotic resis-
tance is interfering with an antibiotic target site either
by preventing the antibiotic to reach its binding site
(target protection) or by target site modification that
decreases an antibiotic binding affinity. Examples of
drugs affected by this mechanisms include tetracy-
cline [Tet(M) and Tet(O)], fluoroquinolones (Qnr),
and fusidic acid (FusB and FusC) resistance (Munita
and Arias 2016). Tet(O) and Tet(M) proteins interact
with the ribosome and dislodge the tetracycline from
its binding site in a GTP-dependent manner, restoring
the protein synthesis (Donhofer et al. 2012; Li et al.
2013). The quinolone resistance protein Qnr belongs
to the pentapeptide repeat protein family and it acts as
a DNA homologue competing for the DNA binding
site of the DNA gyrase and topoisomerase IV. The
reduction in the DNA gyrase–DNA interaction pre-
vents the quinolone molecule from forming the lethal
Phytochem Rev (2019) 18:241–272
DNA–quinolone complex (Aldred et al. 2014). The
antibiotic target site changes may also result from
point mutations in the genes encoding these targets,
enzymatic alterations of the binding sites by methy-
lation, or by ‘‘replacement or bypass of the target site’’.
Classical examples of mutational resistance include
development of rifampin (RIF) resistance (Campbell
et al. 2001) and the resistance to oxazolidinones
(linezolid and tedizolid) (Chen et al. 2013). The
resistance to erythromycin is achieved through the
enzymatic modification of its target site by the
ribosome methylation, which is being catalyzed by
ribosomal methylases (encoded by the Erm genes)
(Leclercq and Courvalin 2002). The ‘‘replacement or
bypass of the target site’’ strategy is used by the
bacteria that are resistant to vancomycin. This antibi-
otic kills bacteria by preventing their cell wall
synthesis by binding to nascent peptidoglycan precur-
sors (D-Ala-D-Ala) and forming a cap that results in the
loss of cross-linking in the polypeptide chain (Gardete
and Tomasz 2014). The vancomycin-resistant bacteria
produce a different variant of the peptidoglycan
precursors (D-alanyl-D-serine or D-alanyl-D-lactate) or
completely destroy the ‘‘normal’’ D-Ala-D-Ala ending
precursors (Hiramatsu 2001; McGuinness et al. 2017).
Occasionally the resistance to antimicrobial agents
can be obtained via combined mechanisms. For
instance, gentamicin resistance, although it does not
rely on the antibiotic modification, it is executed thru
altering the membrane potential and efflux, as well as
16S rRNA methylation (Waglechner and Wright
2017).
Inhibition of bacterial efflux pumps
Bacterial drug efflux pumps can efflux a large number
of structurally unrelated drugs and have a significant
role in the development of antimicrobial resistance in
bacteria (Lubelski et al. 2007). Notably, Wang et al.
(2014) and Lechner et al. (2008b) showed that
biochanin A (20), along with its metabolite genistein
(22) are potentiators of the antibacterial activities of
norfloxacin and berberine in wild-type S. aureus and
M. smegmatis, respectively. However, the inhibitory
effect of those flavonoids on NorA MDR efflux pump
(MFS family) was said to be rather moderate. Mild
inhibitory effects were also reported for the sarothrin
(18) from Alkanna orientalis, which inhibited the
growth of M. smegmatis and S. aureus and possessed
259
NorA efflux pump inhibitory activity. Although the
sarothrin alone is a weak antimicrobial agent, it could
increase the activity of other antimicrobial compounds
by blocking the bacterial efflux pumps (Bame et al.
2013). Furthermore, Fujita et al. (2005) restored the
effectiveness of tetracycline against MRSA, by
baicalein (11)-mediated inhibition of tetracycline
efflux pump (Tet(K)). However, baicalein inhibited
the transport of tetracycline in E. coli KAM32, which
lacks the AcrAB pump. This latter observation
suggests that baicalein inhibits some other extrusion
pump(s) for tetracycline (Fujita et al. 2005). EGCG
(42) also inhibited Tet(K) pumps in staphylococci,
presumably by inhibiting the expression of Tet
proteins (Roccaro et al. 2004).
In contrast to Gram-positive bacteria, Gram-nega-
tive bacteria are resistant to wide range of antibiotics,
mainly due to the low permeability of their cell
membrane. The main mechanism attributed to their
resistance consists of MexAB-OprM and AcrAB-TolC
efflux pumps as well as low porin expression (Brei-
denstein et al. 2011). Daidzein (21), an isoflavone,
showed a very slight modulatory effect on M. smeg-
matis as an efflux pump inhibitor (Lechner et al.
2008b). However, molecular docking calculations and
in vitro assays point it as an inhibitor of the MexAB-
OprM and AcrAB-TolC tripartite efflux pumps exist-
ing in P. aeruginosa and E. coli (Aparna et al. 2014).
Daidzein potentiated the efficacy of carbenicillin and
levofloxacin antibiotics against both E. coli and P.
aeruginosa. Furthermore, authors suggested that
daidzein possibly circumvents the efflux resistance
mechanism. The molecular dynamics studies per-
formed by Suriyanarayanan and Sarojini Santhosh
(2015) reported that quercetin (32) could bind to M.
tuberculosis Mmr and E. coli EmrE efflux pumps,
suggesting that it may downregulate the drug efflux
and thus play a role of non-antibiotic adjuvant. A study
by Dey et al. (2015) examined the antimicrobial
activity of EGCG (42) and quercetin against drug-
resistant M. tuberculosis and b-lactamase producing
K. pneumoniae and demonstrated the antimicrobial
effects of both flavonoids. The results of Kurinčič et al.
(2012) demonstrated that EGCG shows antibacterial
activity and enhances antibiotic effects against clinical
isolates of P. aeruginosa, and EGCG was proposed to
act as an inhibitor of the efflux pump MexAB-OprM
(Kurinčič et al. 2012). Similarly, EGCG, by impairing
CmeDEF drug efflux systems, partially reversed the
123
260
drug resistance of Campylobacter spp, (Kurinčič et al.
2012). Moreover, Christena et al. (2015) showed the
role of efflux pumps in quorum sensing, cell-to-cell
signaling, and biofilm formation.
Altogether, these reports suggest that flavonoids act
rather as efflux pumps potentiators than inhibitors, and
the mechanistic relationship between efflux pumps
and biofilm formation requires further studies. The
need for further studies is highlighted by the fact that
efflux pumps make antibiotics ineffective, and the
combination therapy along with the existing flavonoid
inhibitors could solve this problem.
Antimicrobial action vs ROS production
It must be accepted that the mammalian innate
immune system has evolved with sophisticated mech-
anisms to recognize and kill bacteria. These processes
are mediated mainly by the phagocytosis mechanism,
by which macrophages and neutrophils engulf bacte-
rial cells to kill them by an ‘‘oxidative burst’’ produced
by the NADPH oxidase, a main source for the
generation of ROS in activated neutrophils and
macrophages (Nunes et al. 2013).
A lot of studies have showed that the bactericidal
antibiotics such as b-lactams, aminoglycosides, and
fluoroquinolones induced oxidative stress, regardless
of their specific targets, and participated in the ROS-
antibiotic bacteria killing [reviewed by Dwyer et al.
(2014) and Vatansever et al. (2013)]. On the other
hand, several other reports failed to show the link
between ROS and antibiotic-mediated killing [re-
viewed by Van Acker and Coenye (2017)]. These
inconsistent data may have resulted from the presence
of ROS, which are generated through the hyperacti-
vation of normal cell metabolism, as well as the related
difficulty or even the impossibility to completely
separate the effects of decreased ROS levels and ROS
production as a consequence of the action of antibi-
otics (Dwyer et al. 2014; Van Acker and Coenye
2017). Flavonoids are considered as efficient ROS
scavengers; however, the flavonoid concentration in
human plasma and most tissues is too low to effec-
tively reduce ROS (Brunetti et al. 2013). Furthermore,
flavonoid ROS scavenger usage should be carefully
considered, since low ROS concentrations are, on the
contrary, beneficial for bacteria and can induce
resistance. Thus, the function of flavonoids as an
antimicrobial potentiator should rather be associated
123
Phytochem Rev (2019) 18:241–272
with the regulation of the activities of different
proteins and molecular processes, and there is need
for further studies, especially regarding their syner-
gistic action.
Combined action of flavonoids and antibiotics
As already mentioned above, one of the suggested
approaches for improving the antibiotic efficiency
against bacteria involves the use of flavonoids as
potentiators (Brynildsen et al. 2013). Moreover,
flavonoids are used by cells for their protection against
the harmful effects of ROS (Baldim et al. 2017;
Brunetti et al. 2013; Pietta 2000; Prochazkova et al.
2011). Notably, Brynildsen et al. (2013) proposed to
increase the antibiotic efficacy not by impairing the
organism’s ROS defense systems by adjuvants, such
as flavonoids, but by amplifying the endogenous ROS
production, which should compromise its ability to
cope with an oxidative attack from the antibiotic.
Kohanski et al. (2007) demonstrated that quinolones,
b-lactams, and aminoglycosides stimulated hydroxyl
radical formation via the Fenton reaction. Addition-
ally, both the iron chelator and the hydroxyl radical
quencher, which could be flavonoids, attenuate killing
by bactericidal drugs, which suggest that hydroxyl
radicals contribute to bactericidal antibiotic-mediated
cell death. Furthermore, uptake of aminoglycoside
antibiotics (AGs: gentamycin, amikacin, neomycin,
streptomycin, spectinomycin, and tobramycin) is
driven by the proton motive force (Taber et al.
1987), which is abolished when ROS concentrations
are increased over wild-type levels (Ezraty et al. 2013;
Farha and Brown 2013). Flavonoids like other iron
chelators, protect against AGs by blocking AGs
uptake via the impairment of Fe-S cluster synthesis
resulting in the impendence of the PMF (Ezraty et al.
2013).
The most common mechanism of AG resistance is
the chemical modification by bacterial aminogly-
coside-modifying enzymes (AMEs), acetyltrans-
ferases (AACs), nucleotidyltranferases (ANTs), or
phosphotransferases (APHs) (Ramirez and Tolmasky
2010). Unfortunately, only few flavonoids were
reported as inhibitor of these enzymes. The quercetin
(32) was proposed as an APH inhibitor (Daigle et al.
1999; Shakya et al. 2011) and was shown to occupy the
ATP binding site and to interact with the enzyme
APH(200 )-IVa through a series of hydrogen bonds.
Phytochem Rev (2019) 18:241–272
Moreover, apigenin (10) although did not affect
APH(200 )-IVa, it was able to inhibit the closely related
enzyme APH(200 )-IIa. Furthermore, metal cations
(Mg2?, Cr3?, Cr6?, Mn2?, Co2?, Ni2?, Cu2?, Zn2?,
Cd2?, and Au3?) have been demonstrated to inhibit
the AG acetyltransferase activity and to increase the
efficacy of AGs in resistant strains (Li et al. 2015)
Therefore, flavonoids as chelators could be used as a
potential inhibitors of AMEs. However, such flavo-
noid application requires future research.
It should be considered, that combined use of
antibiotics with flavonoids can lead to some negative
effects. For example, isoquercetin (33) showed antag-
onism with aminoglycoside antibiotics such as
neomycin, kanamycin, gentamicin, and amikacin
when tested with E. coli 27 strain. However, quercetin
did not affect the antibacterial activity of the amino-
glycoside antibiotics. Moreover, both isoquercetin and
quercetin (32) did not affect the action of aminogly-
cosides against a multiresistant strain of S. aureus
(Veras et al. 2011).
Testing antimicrobial activities and reasons
for discrepancies in results
Antimicrobial flavonoids are often described by the
minimum inhibitory concentration (MIC), which is
being their minimum concentration that causes visible
inhibition of bacterial growth. MIC assessment is
usually the first step of evaluation of new antimicro-
bials and it is determined in agar dilution or broth
dilution assays (O’Neill and Chopra 2004). Plant
extracts with MIC B 100 lg/mL and purified com-
pounds with MIC B 10 lg/mL are considered
promising (Rios and Recio 2005). However, MIC
parameter describes the bacteriostatic activity of the
given compound only, same as Kirby-Bauer’s agar
diffusion test, which is also commonly used in the
antimicrobial susceptibility testing (Ahmed et al.
2016; Awouafack et al. 2011; Tohma et al. 2016).
However, with the increasing number of immuno-
compromised patients, it is important to develop a
bactericidal drug, rather than just a bacteriostatic
(Corti et al. 2009). Bactericidal activity is determined
by the minimum bactericidal concentration (MBC) in
time-kill assays. MBC and MIC parameters comple-
ment each other and MBC below the four times MIC
value suggests the bactericidal action of a tested
261
compound (French 2006). However, bactericidal
studies depend on determining the Colony-Forming
Units (CFU) number, while some flavonoids have
been reported to induce the formation of multicellular
aggregates. Thus, decrease in CFU numbers may
result rather from the cell aggregation, than the
bactericidal action of a tested compound. Since
MBC studies of flavonoids are often unreliable, other
assays must be used to ensure the lack of cell
aggregation. Microscopic study-supported time-kill
assays suggested by Cushnie et al. (2007) can be a
solution to this problem.
Furthermore, many other factors may affect the
results of antimicrobial in vitro studies, either it is the
MIC, MBC studies or it is the Kirby-Bauer’s antibiotic
test. The most important variables include the sensi-
tivity of strains, antimicrobial potential of a studied
compound, the type of medium and the optical density
of the inoculum (CLSI 2017). Clinical and Laboratory
Standards Institute (CLSI) is one of the organizations
that standardized many of these variables (CLSI
2017). For example, Müller-Hinton broth/agar is
accepted as a standard growing medium for antimi-
crobial susceptibility testing, while the cell density of
the inoculum for broth microdilution assay is typically
at 5 9 105 (Wiegand et al. 2008). However, not all
research teams follow CLSI guidelines, or even any
guidelines at all. Moreover, there are limitations to
each antimicrobial assay, for example, a flavonoid
with poor agar diffusion abilities will yield weak
results in agar diffusion test, despite its possibly good
antimicrobial activity (Zheng et al. 1996). Further-
more, the solvent used for the preparation may
influence the extract contents and affect the antimi-
crobial activity. Crude methanolic plant extracts
typically have the highest concentration and highest
number of flavonoids, and thereby the strongest
antimicrobial activity (Dar et al. 2016). However,
sometimes the pure compounds are isolated, which are
usually hydrophobic and may precipitate in wrong
solvents, e.g., water (Lof et al. 2011). This will lead to
their reduced contact with the bacterial cells and thus
decreased activity. Notably, some flavonoids have
been known to form salts in alkaline solvents that can
also influence their biological activities (Cushnie et al.
2003). Dimethyl sulfoxide (DMSO) that is typically
used to dissolve isolated flavonoids offers good
polyphenol solubility; however, the DMSO may also
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262 Phytochem Rev (2019) 18:241–272

affect results by interacting with bacterial membranes compounds is typically described by the fractional
(Mi et al. 2016). inhibitory concentration index (FICI) (Wang et al.
2014)
MICðantibiotic aloneÞ
The most potent antimicrobial flavonoids FICI ¼
MICðantibiotic combined with compoundÞ
MICðcompound aloneÞ
In Table 1, we have summarized antimicrobial þ
MICðcompound combined with antibioticÞ
flavonoids with the MIC value below 10 lg/mL. We
chose the MIC value as a determinant, because it is the
Staphylococcus aureus with many resistant strains
most commonly used characteristic of novel antimi-
is one of the most dangerous pathogens nowadays
crobials, including flavonoids. MBC values (if tested)
(Lindsay 2013). Thus, there is an understandable
are also presented in the Table 1. Given the possibility
desire to find agents that would enhance the available
of cell aggregation during MBC studies, as well as
anti-staphylococcal drugs, and studies of most
other potential reasons for results discrepancies, these
research teams are focused on the synergy of
data must be interpreted with caution. In majority of
flavonoids and antibiotics against resistant S. aureus
studies cited in Table 1, mechanisms of antibacterial
strains. Reports of synergy and additive effects of
action of tested flavonoids have not been elucidated.
flavonoids and antibiotics are summarized in Table 2.
Mechanism of action and structure–activity relation-
Although the involvement of flavonoids in the bacte-
ship studies are usually conducted by research teams
rial growth control is extensively studied, their
of different specialty, compared to those who report
complexation with antibiotics remains poorly
novel antimicrobial agents. It is understandable, since
understood.
those areas of studies require different approach and
expertise. It does, however, create a knowledge gap,
where a lot of compounds are known for their
Concluding remarks
antimicrobial activity, but little detail is known about
mechanism of action of every one of them. Some of
Recently, CDC estimated that one in five pathogens
the compounds present in the Table 1 had been studied
from hospital-acquired infections represents mul-
for their mechanism of action in different studies and
tidrug-resistant strain (MDR) (Weiner et al. 2016),
have been described above.
while there is no progress in the development of new
classes of antibiotics. Hence, there is a serious need for
finding new antimicrobial agents or at least substances
Examples of synergy and additive effect
that would enhance the effectiveness of current drugs
between flavonoids and antibiotics
(Abreu et al. 2012). Notably, many flavonoids show
strong antimicrobial effects and/or synergy with
Recently, there has been a growing interest in
‘‘conventional’’ antibiotics. There are also reports of
uncovering novel antibiotic adjuvants through sys-
flavonoids inhibiting bacterial virulence factors, such
tematic approaches. Notably, the ability of plant
as hemolysis activity of S. aureus (Qiu et al. 2010).
metabolites to enhance the activity of antibiotics has
Importantly, most of the flavonoids are considered
been widely reported (Sana et al. 2015). These
nontoxic because of their ubiquity in all sorts of plant-
compounds that have potential activity against patho-
derived foods and beverages. Few toxicity studies
genic bacteria are variably been termed modulating,
support that notion. Dzoyem et al. (2013) conducted
resistance modifying, or reversal adjuvants. In this
experiments on silkworm larvae, which supported low
review, we provide examples of synergy between
or no toxicity of tested flavonoids. Single-dose toxicity
antibiotics and flavonoids. Most of the researchers
studies performed on lab rats also failed to determine
propose flavonoids to be resistance modifying agents
methanolic extract of flavonoids as toxic (Kuete et al.
(RMAs). The mechanisms of RMA action may
2007). Moreover, Ames test showed no mutagenic
include inhibition of efflux pumps or antibiotic-
effect of the selected flavonoids (Bagla et al. 2014).
degrading enzymes and membrane permeabilization
Daily intake of flavonoids is estimated at
(Abreu et al. 2012). Interaction of two antimicrobial

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Table 1 Strongest antimicrobial flavonoids reported in recent S. aureus; NT—not tested; PPSA—penicillinase-producing S.
years (EGCG—epigallocatechin gallate; MRSA—methicillin- aureus; VISA—vancomycin-intermediate S. aureus; VRE—
resistant Staphylococcus aureus; MSSA—methicillin-sensitive vancomycin-resistant Enterococci)
Flavonoid MIC/MBC (lg/mL) Strain References

Flavone 1.95/3.9 P. vulgaris, P. mirabilis Basile et al. (2010)

Isolupalbigenin 1.56–3.13/6.25–25 MRSA Sato et al. (2006)
Galangin 0.89–14.16/ MSSA, MRSA, Pepeljnjak and Kosalec (2004)
1.38–23.44 Enterococcus spp.,
P. aeruginosa
Rutin 8/16 K. pneumoniae Djouossi et al. (2015)
Rhamnoisorobin 1-2/NT S. aureus, P. aeruginosa, Tatsimo et al. (2012)
S. typhi
2-hydroxylupinifolinol 2.3–4.7/NT MSSA, MRSA, Thongnest et al. (2013)
S. pyrogenes, B. cereus
30 -O-methyldiplacol 2–4/NT B. cereus, E. faecalis, Smejkal et al. (2008)
L. monocytogenes,
S. aureus, S. epidermidis
2,8-diprenyleriodictyol 0.5–4/NT MSSA, MRSA Dzoyem et al. (2013)
Diplacone 2–16/4.9–39.2 MRSA Navratilova et al. (2016)
Hesperetin 4–32/NT S. aureus Lopes et al. (2017)
Naringenin C 2.8/NT M. tuberculosis Chen et al. (2010)
Pinocembrin 3.5/NT M. tuberculosis Chou et al. (2011)
Sepicanin A 2.9/2.9 MRSA Radwan et al. (2009)
Dihydrokaempferol 6.25/12.5–25 VRE, S. aureus Tajuddeen et al. (2014)
Bartericin A 0.31–0.61/NT C. freundii, S. dysenteriae, Kuete et al. (2007)
B. cereus, S. aureus,
S. faecalis (among others)
Isobavachalcone 0.3–0.6/0.6–1.2 S. faecalis, S. aureus, Mbaveng et al. (2008)
E. aerogenes, E. cloacae
(among others)
Panduratin A 1–2/4–8 E. faecalis, E. faecium Rukayadi et al. (2010)
Phloretin 1/NT S. aureus Lopes et al. (2017)
Licochalcone A 2-8/NT MSSA, MRSA Qiu et al. (2010)

100–1000 mg/day, depending on the diet (Aherne and
O’Brien 2002). In general, no adverse effects have
been associated with high dietary intakes of flavonoids
from plant-based food. Flavonoid-rich foods and
beverages include tea, red wine, fruit skins, citrus
fruits, berry fruits, and honey (Kumar and Pandey
2013). Those foods are typically attributed to many
health benefits. The lack of toxicity and natural
occurrence makes flavonoids possibly good food
preservatives. They can be a viable candidate for
replacing synthetic preservatives that are disliked by
the consumers (Wu et al. 2013). The lack of adverse
effects may be explained by the relatively low
bioavailability and rapid metabolism that leads to
elimination of most of the flavonoids (Harwood et al.
2007; Ottaviani et al. 2015). To date, the importance of
the safe use of flavonoid supplements in pregnancy
and lactation has not been well established (Hendler
and Rorvik 2009; Mills et al. 2013). Moreover, the use
of green tea extracts was directly associated with
abnormally high levels of liver enzymes (Dostal et al.
2015; Sarma et al. 2008). Obviously, further toxicity
studies are needed before releasing any food or
medicine containing high amounts of flavonoids.

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264 Phytochem Rev (2019) 18:241–272

Table 2 Examples of synergy and additives effect between antibiotics and flavonoids (FICI—fractional inhibitory concentration
index; EGCG—epigallocatechin gallate; MSSA—methicillin-sensitive Staphylococcus aureus; MRSA—methicillin-resistant S.
aureus; PPSA—penicillinase-producing S. aureus, VISA—vancomycin-intermediate S. aureus)
Flavonoid Antibiotic FICIa Strain References

Flavone Vancomycin 0.096 VISA ATCC 700699 Bakar et al. (2012)

Oxacillin 0.126
Apigenin Ampicillin, 0.18–0.47 MRSA strains Akilandeswari and Ruckmani
Ceftriaxone (2016)
Baicalein Tetracycline 0.06–0.12 MRSA strains Fujita et al. (2005)
Baicalein Penicillin 0.14–0.25 PPSA strains Qian et al. (2015)
Amoxicillin 0.14–0.38
Baicalein Cloxacillin \ 0.02 S. aureus DMST 20651 Eumkeb et al. (2010)
Diosmetin Streptomycin 0.39 S. aureus 1199B, RN4220 Wang et al. (2014)
Ciprofloxacin 0.09 S. aureus EMRSA-15
Luteolin Ampicillin, 0.82–0.9 MRSA ATCC 43300 Usman Amin et al. (2016)
Cephradine,
Ceftriaxone,
Imipenem,
Methicillin
Luteolin Ceftazidime 0.37 S. pyogenes DMST Siriwong et al. (2015)
Genistein 0.27 30653 - 30655
Genistein Norfloxacin 0.38 S. aureus 1199B, Wang et al. (2014)
RN4220,
Ciprofloxacin 0.09 S. aureus EMRSA-15
Galangin Cloxacillin \ 0.02 S. aureus DMST 20651 Eumkeb et al. (2010)
Morin Ampicillin 0.31 MRSA ATCC 3359 Mun et al. (2015)
0.75 MRSA DPS-1
Myricetin Isoniazid 0.2 M. smegmatis mc2155 Lechner et al. (2008a)
Galangin Amoxicillin \ 0.09 E. coli (AREC) Eumkeb et al. (2012)
Kaempferide
Kaempferide-3-O-
glucoside
Quercetin Cloxacillin \ 0.02 S. aureus DMST 20651 Eumkeb et al. (2010)
Quercetin Ceftriaxone, 0.66–0.84 MRSA ATCC 43300, Usman Amin et al. (2016)
Quercetin ? luteolin Imipenem, 0.45–0.65 MRSA Clinical Isolates
Rutin ? morin Methicillin 0.8–0.9
EGCG Tetracycline 0.375 MRSA6975, MRSA3202 Navratilova et al. (2016)
Oxacillin 0.5
Synthetic Vancomycin 0.97 E. faecium Budzynska et al. (2011)
3-arylideneflavanones Oxacillin 0.01–0.58 S. aureus A3
a
‘Synergy’ was defined where the FICI was less than or equal to 0.5; whilst ‘additive’ effects were observed when the FICI was
greater than 0.5 and less than or equal to 1.0; greater than 1 and less than 2 as indifferent; Antagonistic effects were observed when
the FICI was greater than 2.0

Furthermore, to increase the specificity and safety of Considering the hydrophobic nature of flavonoids,
flavonoids more focus on their mechanisms of action few questions are raised regarding their in vivo
and a structure–activity relationship is required. activity, like ‘‘how to achieve and sustain their high

123
Phytochem Rev (2019) 18:241–272
blood serum concentration?’’ Their structural modifi-
cations or use of drug carriers may be essential to
modulate their infiltration into the bloodstream. On the
other hand, getting to know flavonoid metabolism in
mammalian cells may be helpful in preventing their
rapid catabolism. Thus, determining if flavonoids are
effective antimicrobials at in vivo environment
remains crucial. Finally, flavonoids maintain their
biological activity, thanks to a finely regulated trans-
port and accumulation system that allow entrance into
different subcellular compartments. Nevertheless, a
comprehensive view of the phenomenon has not yet
been proposed and is still under investigation.
Compliance with ethical standards
Conflict of interest The authors have declared that there is no
conflict of interest.
Open Access This article is distributed under the terms of the
Creative Commons Attribution 4.0 International License (http://
creativecommons.org/licenses/by/4.0/), which permits unrest-
ricted use, distribution, and reproduction in any medium, pro-
vided you give appropriate credit to the original author(s) and
the source, provide a link to the Creative Commons license, and
indicate if changes were made.
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